The overall objectives of the proposed studies are two-fold. The first is to determine the physiological control over cumulus expansion and GAG synthesis. To date, this control appears to be manifest primarily in the actions of gonadotropins. However, the roles of intrafollicular components has not been examined. The second objective is to determine the roles of intracellular components and structures in these processes. The specific aims of the proposed studies are to determine: 1. the effects of follicle size on gonadotropin induced cumulus expansion and glycosaminoglycan (GAG) synthesis, 2. the sensitivity of cumulus-oocyte complexes to FSH and LH induced expansion and GAG synthesis, 3. the effects of follicular fluid on cumulus expansion and GAG production, 4. the effects of guanosine cyclic monophosphate on expansion and GAG production, 5. if Ca++ and/or calmodulin are involved in expansion and GAG synthesis, and 6. if the cytoskeleton is involved in the expansion process. In all cases of morphological investigation (expansion), cumulus expansion will be evaluated by microscopic examination. In all cases of biochemical investigation (GAG synthesis), 3H-glucosamine incorporation into GAG will be determined by HPLC separation and liquid scintillation counting. Morphological and biochemical evaluations will be made on the same cumulus-oocyte complexes after culture. Responses of the complexes from follicles at progressive stages of development to gonadotropins and follicular fluid will be determined qualitatively and quantitatively. The involvement of calmodulin and the cystoskeleton in expansion and GAG synthesis will be determined with appropriate blocking agents. The role of Ca++ in expansion and GAG synthesis will be determined by the use of a Ca++ free culture medium.